This invention relates generally to the field of G protein coupled receptor (GPCR) expression and modulation.
G protein-coupled receptors (GPCRs) are a historically successful therapeutic target family, with GPCR-directed drugs covering a wide range of therapeutic indications. As cell surface receptors, GPCRs are vital to cellular functioning, because they are primary mediators of cell to cell communication.
Mammalian cells express very low levels of endogenous GPCRs, generally with no more than three thousand copies per cell. This level is sufficient for receptor function, but offers a challenge to GPCR research, which often requires much higher concentrations of functional proteins. For example, structural studies, small molecule drug design and generation of functional antibodies against the native GPCR conformation require expression levels that are orders of magnitude higher than what is seen using current methods.
Attempts have been made to isolate mammalian cell lines that overexpress exogenous GPCRs, but these past attempts have failed due to the cellular toxicity that occurs with receptor overexpression. Attempts to create expression systems in “lower” organisms have similarly met with limited success due to inefficient folding (bacteria), low yield (yeast) or incorrect post-translation modification (baculovirus).
A need thus exists for stable, high-expression systems capable of providing multiple copies of GPCR proteins for structural and functional studies.